New Insights From Cardiac Muscle Applied To Skeletal Muscle

2y ago
19 Views
2 Downloads
442.75 KB
7 Pages
Last View : 18d ago
Last Download : 2m ago
Upload by : Victor Nelms
Transcription

Review ArticleMore InformationNew insights from cardiac muscleapplied to skeletal muscle*Address for Correspondence: Gerry A Smith,University of Cambridge, Retired, 14, LantreeCrescent, Trumpington, Cambridge CB2 9NJ,UK, Tel: 01223 515394;Email: gas1000@cam.ac.ukGerry A Smith*Retired, University of Cambridge, Cambridge CB2 9NJ, UKIntroductionI have recently described the origin of the second Ca2 binding in the triggering of contractile activity in cardiacmyo ibrils that is the origin of the Ca2 Hill coef icient of 2for the ATPase. This site is not a simple protein binding siteand cannot be measured by 45Ca2 binding. The myo ibrilprotein unit requirements are described by me and so are theconsequences of disruption of the function of these units andthe related medical outcomes. The purpose of this paper is toreview the topic and extend the reasoning to the function ofskeletal muscle and cite the literature that supports this.Submitted: November 10, 2020Approved: January 14, 2021Published: January 15, 2021How to cite this article: Smith GA. New insightsfrom cardiac muscle applied to skeletal muscle. JCardiol Cardiovasc Med. 2021; 6: 007-013.DOI: 10.29328/journal.jccm.1001109Copyright: 2021 Smith GA. This is an openaccess article distributed under the CreativeCommons Attribution License, which permitsunrestricted use, distribution, and reproductionin any medium, provided the original work isproperly cited.Abbreviations: MyBP-C: Myosin BindingProtein-C (was called protein-C); TNN: GeneName in Database; TnC: Troponin-C; TnI:Troponin-I; Tm: Tropomyosin; TnT: Troponin T;pCa50: The point with 50% activationSummaryIn muscle activation Ca2 binds to the activating subunittroponin-C (TnC) which changes its interaction with anothersubunit troponin-I (TnI) which changes the interaction of thiswith yet another subunit tropomyosin (Tm). The consequenceof this complex activity is the unblocking of interaction ofthe myosin light chain (LC), ATP bound, with the actin chainallowing the formation of a cross-bridge between the thickand thin ilaments and the resulting ATPase activity givingmuscle contraction and/or force. The cycle is completed byreplacement of the ATPase products (ADP and phosphate Pi)by the binding of MgATP to the myosin light chain. This is thestory accepted to date in all striated muscle systems. Howeverin the cardiac system I irst suggested in 2001 [1] that thiswas not all embracing and mooted that a further Ca2 bindingwas required to satisfy the observed cooperativity (raisedHill coef icient for Ca2 activation 1). This I have recentlyrevisited [2] to show the implication of yet another myo ibrilsubunit in the Ca2 activation of myocyte contraction, i.e. themyosin binding protein-C (MyBP-C) [2-4] and applied thisto the cardiac system [5], and thus demonstrating the originof many cardiomyopathies, particularly hypertropy [5]. Thekey to these results is that the MgATP bound myosin is thefully relaxed state, it is not used in the functioning of thecross-bridge and in the unperturbed system the presence ofintact cMyBP-C ensures this. There are many references to thecMyBP-C acting to modulate the Ca2 sensitivity of the 09OPEN ACCESSmuscle but absolutely none give clear mechanistic explanationof how this occurs. My recent papers show that along withthe cTnI the cMyBP-C ensure the Myosin bound MgATP hasto become myosin bound CaATP before the cross-bridge canform. In the heart this binding of Ca2 requires a concentration[Ca2 ] well above that required by the TnC binding and is alsoinhibited competitively by Mg2 . Any failure of this concertedaction of the subunits, cTnI and cMyBP-C, results in the bindingof Ca2 by cTnC becoming the sole requirement for activationof contraction, this produces some contraction at much lower[Ca2 ] and hence incomplete relaxation in diastole. The resultof incomplete relaxation in diastole is chronic tension that istransmitted to another myo ibril component, the giant proteintitin, this tension acts at the spring region of the titin releasinga myocyte growth factor the result of which is unwantedgrowth, i.e. hypertrophy.For considering extending the above indings to skeletalmyo ibrils the key lead in citations involve those that depletethe sarcomere of subunits by the use of EDTA to remove allbound divalent cations from their structural binding sites.This also includes the substitution with differently resourcedunits or retention of speci ic proteins [6,7]. I initially layhttps://www.heighpubs.org/jccm007

New insights from cardiac muscle applied to skeletal musclethe ground by giving a comprehensive review of the threesubunits intimately involved.BackgroundIn this study I concentrate on three proteins that areessential components of the Ca2 stimulated control of musclecontraction. The proteins are the Ca2 binding activating unitTroponin-C (TnC), the inbibitory unit Troponin-I (TnI) andmyosin binding protein-C (MyBP-C), table 1. These all showisoforms speci ic to adult muscle type, fast and slow skeletaland cardiac. The gene name and number for TnI and MyBP-Care in agreement, three genetic versions slow skeletal, fastskeletal and cardiac but only two, fast skeletal and slow/cardiac for TnC.The notable differences between these protein isoformsTnI comparisons: The most obvious difference arebetween the cardiac and the other (skeletal) versions. Thecardiac has an extended N terminal sequence that folds andbinds to the inhibitory region (residues 137-148) Figure 1.Layland, et al. [9] report “binding of Ca2 to cTnC duringsystole induces conformational changes that relieve theinhibitory in luence of cardiac cTnI”. More correctly put thisshould read “binding of Ca2 to troponin C and myosin ATP .”Phosphorylation of cTnI alters its function, in particularthe threonine of the cardiac version (residue 142 or 3or 4 depending on the reference used) which undergoesTable 1: The isoforms of the key sarcomere subunits (from Uniprot).Protein isoformsSlow skeletalmuscle ssTnIFast Skeletalmuscle fsTnICardiac .519q13.4Protein isoformsSlow SkeletalMuscle ssMyBP-CFast SkeletalMuscle fsMyBP-CCardiac .219q13.3311p11.2Protein isoformsSlow SkeletalMuscle ssTnCFast SkeletalMuscle fsTnCCardiac .123p21.1phosphorylation by PKC-βII under diastolic stretch, theFrank-Starling law [3,4,10], all the TnIs also get serinephosphorylated in various places altering function.Mutations of TnITo date, no human disease has been reported withmutations in ssTnI [11]. However mutations in the fsTnI genehave been found to cause myopathy and distal arthrogryposis(DA). A missense mutation R174Q, a nonsense mutation(premature stop codon R156X), and three in-frame deletionmutations ΔE167, ΔK175 and ΔK176 have been reported inDA patients [12]. The mutations associated with DA are allin the C-terminal actin-tropomyosin binding domain. Manymutations of cTnI have been found to cause hyper, restrictive,and dilated myopathies [5,13,14].MyBP-C comparisonsThe name, now changed from protein-C to myosin bindingprotein-C arises from the irst indings that it binds to theS2 region of myosin, Figure 2. The last realised unit of thesarcomere can only now be recognised as important as TnCin triggering of contraction, if not more so. It is unfortunatethat the binding to actin has been so overlooked in manystudies. The FHL-1 gene binding couple with the titin bindingis probably re lected in myo ibril growth, hypertrophy, as isthe FHOD-3 binding.TnC comparisonsA comprehensive review by Katrukha [8] has manyleading references within (Figure 3). All TnC forms have fourCa2 -binding EF-hands that are combined pairwise into theN-terminal (sites I and II) and C-terminal (sites III and IV)globular domains. In the fast skeletal isoform of TnC, all fourEF-hands are able to bind Ca2 or Mg2 . Only the N-terminaldomain EF-hands I and II have a lower af inity but highselectivity for Ca2 and both play a crucial role in the regulationNote, the ssTnC and cTnC are the same protein.Figure 1: Domain structure of human cTnI [8]. a) Scheme of the cTnI secondarystructure. The N-terminal domain of TnI is residues 2-32, the α-helices H1-H4 in theTnI, and the mobile domain H4-H6. The wavy curve represents the Xaa-Pro regionof cTnI (residues 12-18) forming a proline helix. Short β-strands 1and 2 are markedby arrows. Proteins of the thin filament that interact with the relevant regions ofthe cTnI molecule are indicated as ovals. N-TnC and C-TnC, N- and C-terminaldomains of TnC, respectively; Tm, tropomyosin. b) Domain organization of cTnI.ID, inhibitory domain; RD, regulatory 09Figure 2: Schematic diagram of full length slow skeletal (ss), fast skeletal (fs) andcardiac (c) MyBPC paralogs. Each isoform comprises three Fn3 domains andseven or eight Ig domains. The known binding partners and positions are indicatedby the horizontal stripes below. Note the regulatory phosphorylation sites in theP/A and M domain of the ssMyBP-C and cMyBP-C paralogs are indicated bysmall black ellipses. The cMyBP-C has an additional 28 amino acid loop in the C5domain (15,) and one clearly obviously wrong reference [17].https://www.heighpubs.org/jccm008

New insights from cardiac muscle applied to skeletal muscleFigure 3: Domain structure of human cardiac/slow skeletal cTnC. The α-HelicesN and A-H are represented by cylinders, β-strands by arrows. Two circles withoutdesignation mark the structural high affinity sites III and IV. The circle designatedwith “Ca” represents the one high-specificity region that binds Ca2 , Site II. Proteinsof the thin filament that interact with the relevant regions of the troponin-T (TnT)molecule are indicated in ovals. N-TnI, N-terminal domain of TnI; RD, regulatorydomain; ID, inhibitory domain.of muscle contraction. Sites III and VI are high af inity, are ionbound under all physiological conditions and structural andbind TnC to the sarcomere.A few amino acid substitutions in the irst EF-hand ofthe c/ssTnC isoform (site I) impede the ion binding so thatc/ssTnC has only three Ca2 -binding sites, only one active incontraction stimulation.The activation binding of Ca2 to the two types of TnC iswell de ined by Lee [17] and has been resolved for some time[18-20]. In the heart the single activating Ca2 binding site incTnC gives rise to a Ca2 Hill coef icient of 2 when measuredcarefully on the completely intact system. This cooperativityof a second site with the single TnC site II is explained asarising from an exchange of Mg2 for Ca2 on the myosin lightchain controlled by cMyBP-C [1-4]. For the fast skeletal musclethe situation is more muddled as the Hill coef icient, especiallywhen measuring tension can be much higher than the valueof 3 that is anticipated if the two fsTnC sites work with thefsMyBP-C in the same way as cardiac [21].The sarcolema complexScheme 1, the cross-bridge cycleThe ATPase protein complex consists of MyBP-C whichthrough its N-terminus (S1S2) binds both thick ilament(Myosin) and thin ilament (Actin). Creatine Kinase is boundto the MyBP-C. The MyBP-C S1S2 fragment when added bindsto the actin and myosin, scheme 1, in the same way as thewhole in the spaces between the MyBP-C units. The fragmentdoes not have the MgATP inhibitory function of the whole andbypasses the Ca2 swap for Mg2 as does the MyBP-C removal,this cation swap obeys normal competitive binding. Theilaments are associated along length with the giant proteinTitin which is structural and reacts to chronic stress with therelease of sarcolemal growth factors.Results and discussionIn the light of my previous studies on the cardiac system[1-5], in particular the effect of removal of cTnC, Figure 4 [7],my start point was the same treatment given earlier to skeletalmuscle by Brandt, et al. [6]. The data quoted are nearly allpresented as fractional tension v pCa charts the shifts andslope changes of which are informative of what is happening.The extraction of cTnCFirst I quote from Hofmann the results obtained withthe cardiac system, Figure 4 (and later Figures 8 and 9 forcomparison with their other data), I follow with the additionof an N-terminal fragment of MyBP-C to cardiac myo ibrils,Figure 5, to introduce the MyBP-C extraction problem withother data and The Brandt data giving igure 6.The addition of the N-terminal fragment of MyBP-C,C1mC2, to cardiac myofilamentsThe addition of the N-terminal fragments of MyBP-C,Figure 5 [22] to the cardiac myo ibrils changes the responseof the system activation to the level of [Ca2 ] de ining theimportance of MyBP-C in the control mechanism.Scheme 1: The cross-bridge cycle.The Actin ilament carries the troponin complex, TnCtroponin-C (the Ca2 binding activation), TnI (the inhibitoryunit)whose inhibitory function is relieved on Ca2 activationand Tm (the tropomyosin) which is moved away to allow theMyLC-Actin cross-bridge to form and last, the TnT troponin ure 4: The effects of cTnC extraction on cardiac myofibrils [7]. Data from amyocyte before cTnC extraction ( ), after partial extraction of cTnC ( ), and finally,with readdition of cTnC ( ). After extraction, tension at pCa 4.5 was 0.49 of controlPo; with readdition, maximum tension returned to 0.75 of control Po.https://www.heighpubs.org/jccm009

New insights from cardiac muscle applied to skeletal muscleFigure 5: Calcium dependence of force in the absence ( ) and the presence ( ) of2μmol/L N-terminal fragment C1mC2 [22].Figure 8: Tension-pCa data obtained before ( ) and after ( ) partial extractionof cMyBP-C from rat ventricular myocytes (with added cTnC). Symbols and errorbars indicate means SEM from 17 myocytes. The data are plotted as relativetension-pCa relationships.Figure 9: Reversibility of cMyBP-C extraction as above.Figure 6: The filled circles ( ) are the % TnC remaining, the open circles ( ) thecalculated Hill coefficient nH which asymptotes to a value of 2.I now realise how the addition of N-terminal fragments givethis result. It was initially believed that the MyBP-C binding tothe myosin S2 region [23] was being displaced by the fragment,this is not true as the Km for the Ca2 independent activation ismuch higher than the concentration used. In fact the MyBP-Cbinds to actin as well as myosin [24] and is located along theilaments spaced out with many more binding sites on bothilaments unoccupied. The N-terminal fragment, especiallywith cardiac ilaments, binds both the actin and the myosin[25], with the many unoccupied binding sites the fragmentbinds to these and as it lacks the MgATP blocking of the wholesubunit it allows the ATPase to function without the Mg2 -Ca2 exchange. The curve is shifted to lower [Ca2 ], towards the Kmfor TnC, and the cooperativity is lost, the Hill coef icient goesto 1.The extraction of fsTnC from skeletal myofibrilsFigure 7: Plots of the dependence of tension on pCa before ( ) and after ( ) partialextraction of TnC tension at each pCa is expressed as a fraction of the tensiondeveloped by the same fiber at pCa 5.0. Error bars indicate SD.https://doi.org/10.29328/journal.jccm.1001109In this early study the ion binding to the structural sitesof TnC is removed by the indiscriminate binding of Mg2 andCa2 by the low selectivity chelator EDTA [6]. This allows thedissociation of the unit from the sarcomere and its washingaway. Doing stopped assays over a time course gave the datahttps://www.heighpubs.org/jccm010

New insights from cardiac muscle applied to skeletal musclein igure 6 and table 2. Note the tension developed greatlyreduces with extraction time limiting the observationspossible to the earlier times.This data from Brandt, et al. [6] is somewhat confusinguntil one realises that the EDTA wash is also removingfsMyBP-C, almost certainly faster than the fsTnC. The resultis the cooperativity, Hill coef icient, reaches that expected forsolely the two Ca2 binding sites on fsTnC being required foractivation. This was 1984 as opposed to reference 7 whichwas later 1991 when this was realised by Moss [7].Partial extraction of skeletal TnC in the presence of free Mg2 By the addition of 1mM Mg2 to the EDTA extractionprocedure the loss of MyBP-C is minimised and this is ensuredby only partial extraction of the fsTnC, Moss, et al. [26]. Note,Figure 7, only a small loss of cooperativity but considerableshift to lower pCa, higher [Ca2 ] i.e. towards the Km of the Mg2 dependent Ca2 activation.Total extraction of TnC from both skeletal and cardiacmyofibrils with reversalMorimoto and Ohtsuki changed the extraction in a similarway to Moss but with a different chelator. Endogenous troponinC in skinned psoas ibers and trabeculae was extracted by CDTAtreatment until the preparations developed no Ca2 activatedtension or ATPase, i.e. complete extraction of TnC. The CDTAtreated preparations were either reconstituted or substitutedwith isolated fast-twitch skeletal or cardiac troponin C. Whenadded back the Hill coef icients are as expected with partialMyBP-C extraction occurring as well. Can this be repeatedwith complete removal of the MyBP-C to give the skeletal andcardiac hill coef icients of 2 and 1 corresponding to fsTnC andcTnC binding alone? (Table 3).Similar extraction results were obtained by Zot and Potter [28].Table 2Minutes extracted02510203031 218 2ParameterTnC (%)100 477 369 840 6Tension (%)100 876 840 628 66.3 0.53.3 0.22.1 0.012 0.3nHThe extraction of cMyBP-C from cardiac myofibrilsI now return to the data of Hofmann and their studies withpartial extraction of subunits of the sarcomere [7]. In this irstof their studies the extraction was limited to removal of solelyMyBP-C by the addition of TnC to the extraction ensuring itremains in the myo ibril. Other damage to the system wasminimised by limiting the degree of extraction igure 8. As theextractions were only partial, unlike the unintentional lossof MyBP-C by Brandt, et al. [6] the precise values of the Hillcoef icients are not relevant, so they are omitted, althoughfollow the expected. The pCa dependency slopes and shiftsare good enough indicators. In this exercise one can easily seethe loss of Ca2 cooperativity with a large shift toward the Kmfor TnC.The process is reversible, Figure 9. Therefore the effect isdue to cMyBP-C.The extraction of fsMyBP-C from skeletal myofibrils(psoas fibres)The same exercise as above was performed on skeletalmuscle myo ibrils igure 10. The authors explain dif iculties inobtaining all the data on reversability. However, although theresults are less spectacular than those from cardiac myo ibrilsthe same effects are observed i.e. a pCa50 shift to lower [Ca2 ]and decrease in the slope.The extraction process was again reversible igure 11.The partial extraction of cTnC, and the subsequentextraction of cMyBP-CI return to the cardiac data reported by Hofmann [7] toemphasise the strength of my earlier argument that thereare two bindings of Ca2 required for activation of musclecontraction, only one of which, on TnC, is measurable bybinding of 45Ca2 . Figure 12 clearly shows the results of eachextraction, both reduce the cooperativity and spectacularlygive pCa shifts in opposite directions towards the Km of that5.97 0.02 5.65 0.03 5.38 0.05 5.21 0.08pCa50Number fibres1113128Table 3Muscle preparationA)B)pCa-tensionpCa .85.72.05.75 skeletal TnC2.75.853.05.94 cardiac TnC1.45.371.55.50 skeletal TnC2.75.113.25.39 cardiac TnC1.55.421.75.74Intact muscleCDTA-treated muscleSkeletalCardiacSimilar extraction results were obtained by Zot and Potter Figure 10: The effects of fsMyBP-C partial extraction on tension developed byrabbit psoas fibers, cumulative data.https://www.

New insights from cardiac muscle applied to skeletal muscle Gerry A Smith* Retired, University of Cambridge, Cambridge CB2 9NJ, UK More Information . and cannot be measured by 45Ca2 binding. The myoibril protein unit requirements are described by me and so are the

Related Documents:

There are three types of muscle tissue: Skeletal muscle—Skeletal muscle tissue moves the body by pulling on bones of the skeleton. Cardiac muscle—Cardiac muscle tissue pushes blood through the arteries and veins of the circulatory system. Smooth muscle—Smooth muscle tis

Mastering a strict bar muscle up will transfer directly to the strict ring muscle ups. There are also other muscle up variations, such as wide ring muscle ups, L-sit muscle ups, weighted muscle ups, explosive muscle ups, one arm muscle ups, etc. This guide is about learning your first

The Motor Unit and Muscle Action Lu Chen, Ph.D. MCB, UC Berkeley 2 Three types of muscles Smooth muscle: internal actions such as peristalsis and blood flow. Cardiac muscle: pumping blood. Skeletal muscle: moving bones. A motor unit consists of a motor neuron and the muscle fibers

between cytoskeletal and contractile elements. Key words: Cytoskeleton, Muscle, Skeletal, Cardiac, Smooth Introduction The muscle cell cytoskeleton has frequently been considered to include those components of the muscle Offprint r

20.1 Types of Movement 20.2 Muscle 20.3 Skeletal System 20.4 Joints 20.5 Disorders of Muscular and Skeletal System . cardiac muscle. Based on appearance, cardiac muscles are striated. They . Fascicle (muscle bundle) Muscle

HASPI Medical Anatomy & Physiology 04c Activity Muscle Tissue The cells of muscle tissue are extremely long and contain protein fibers capable of contracting to provide movement. The bulk of muscle tissue is made up of two proteins: myosin and actin. These

the multiple muscle system of this cockroach leg were found to function differently, where one muscle functions like a motor (muscle 177c) and the other muscle functions like a brake (muscle 179) under in vivorunning conditions (Ahn and Full, 2002). Although bo

Part 1 – Day Trading Explained At DayTradeToWin.com, we mainly focus on one type of market: futures. Some people like to trade stocks, but not everyone has 20,000 to do so. Some people like to trade forex (also called currencies), but not everyone likes the lack of regulation and other shady things in that industry. We prefer to trade futures because they are regulated, are much more .